Structure Of Allergens Research Articles

Differences in structure, allergenic protein content and pectate lyase enzyme activity of some Cupressaceae pollen

Abstract Objective Cupressaceae pollen has commonly been reported to be an important aeroallergen and causal factor of spring, autumn and winter pollinosis in many countries. The aim of this study was to compare of the structure and allergenic protein content of Cupressus arizonica Greene., Cupressus sempervirens L. and Juniperus oxycedrus L. pollen in detail and contribute to Cupressaceae pollen allergen diagnosis and therapy studies in Turkey. Methods The pollen structure were examined by LM and SEM. Pollen protein content was investigated by Bradford protein assay, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), Western blot analysis and two-dimensional polyacrylamide gel electrophoresis (2DE PAGE), respectively. Pectate lyase (PL) enzyme activities were compared. Immunoblotting was carried out by using extracts of the three taxa pollen collected from Turkey. Results All three taxa was found very similar in terms of pollen morphology however, intine thickness was prominently different. Cupressus arizonica pollen extracts showed the lowest PL activity. Five sera specific IgE of all allergic subjects showed reaction with only C. arizonica pollen extracts. Conclusions As a conclusion, the pollen structure, protein function or protein structure and isoforms of allergens could affects allergenic properties of the pollen. This study also may help to improve the Cupressaceae pollen allergen diagnosis and therapy.

Rapid and comprehensive discovery of unreported shellfish allergens using large-scale transcriptomic and proteomic resources

[Extract] Discovery of allergens in various food and inhalant sources is central to our understanding of the molecular mechanisms of allergic reactions. Allergen characterization is the most important underlying factor for better patient management and the design and development of novel immunotherapeutics.1 Of the "Big eight" allergen food groups, shellfish presents a unique challenge in terms of allergen discovery due to the large number and diversity of consumed species, leading to heterogeneity of allergen structure and cross-reactivity among various sources. At present, 31 crustacean allergens have been officially registered in the World Health Organization and International Union of Immunological Societies Allergen Nomenclature Database as compared with 4 mollusk allergens due to pitfalls in current allergy discovery approaches. Cosensitization of patients with crustacean and mollusk allergy is often described; however, the current diagnostic approaches to manage these patients are not based on sufficient molecular knowledge of these shellfish allergens.

Current (Food) Allergenic Risk Assessment: Is It Fit for Novel Foods? Status Quo and Identification of Gaps.

Food allergies are recognized as a global health concern. In order to protect allergic consumers from severe symptoms, allergenic risk assessment for well‐known foods and foods containing genetically modified ingredients is installed. However, population is steadily growing and there is a rising need to provide adequate protein‐based foods, including novel sources, not yet used for human consumption. In this context safety issues such as a potential increased allergenic risk need to be assessed before marketing novel food sources. Therefore, the established allergenic risk assessment for genetically modified organisms needs to be re‐evaluated for its applicability for risk assessment of novel food proteins. Two different scenarios of allergic sensitization have to be assessed. The first scenario is the presence of already known allergenic structures in novel foods. For this, a comparative assessment can be performed and the range of cross‐reactivity can be explored, while in the second scenario allergic reactions are observed toward so far novel allergenic structures and no reference material is available. This review summarizes the current analytical methods for allergenic risk assessment, highlighting the strengths and limitations of each method and discussing the gaps in this assessment that need to be addressed in the near future.

Bovine Milk Allergens: A Comprehensive Review.

Cow milk allergy is one of the most common food allergies in early childhood and often persists through adult life, forcing an individual to a complete elimination diet. Milk proteins are present in uncounted food products, such as cheese, yogurt, or bakery item, exposing allergic persons to a constant threat. Many efforts have been made to overcome this global problem and to improve the life quality of allergic individuals. First, proper and reliable food labeling is fundamental for consumers, but the verification of its compliance is also needed, which should rely on accurate and sensitive analytical methods to detect milk allergens in processed foods. At the same time, strategies to reduce milk allergenicity, such as immunotherapy or the use of food processing techniques to modify allergen structure, have to be extensively studied. Recent research findings on the applicability of food processing, such as heat treatment, fermentation, or high pressure, have revealed great potential in reducing milk allergenicity. In this review, significant research advances on cow milk allergy are explored, focusing on prevalence, diagnosis, and therapy. Molecular characterization of cow milk allergens and cross-reactivity with other nonbovine milk species are described, as well as the effects of processing, food matrix, and digestibility on milk allergenicity. Additionally, analytical methods for the detection of milk allergens in food are described, from immunoassays and mass spectrometry methods for protein analysis to real-time polymerase chain reaction for DNA analysis.

Effects of heating, autoclaving and ultra-high pressure on the solubility, immunoreactivity and structure of major allergens in egg

ABSTRACT Ovotransferrin (OVT) and ovomucoid (OVM) were treated, respectively by different food processing conditions. The protein recovery and immunoreactivity of OVT significantly decreased during heat treatment (P < .05), and aggregates appeared after dry heating at 150°C or 200°C or boiling at 100°C. OVM appeared more stable than OVT under the same treatment. Greatly reduced solubility occurred only after dry heating at 200°C. The immunoreactivity increased after dry heating at 100°C or 150°C or moist heating (60°C, boiling). Autoclave sterilization had a strong effect on the characteristics of both OVT and OVM. Ultra-high pressure had very little effect on OVT and OVM, except at 600 MPa in the moist treatment. Results showed that temperature synergistic processing with water and high pressure had the greatest impact on the characteristics, which could be a potential strategy for mitigating the negative effects of allergens in egg.

Detection of Allergen-IgE interaction in Allergic Children Through Combined Impedance and ROS Measurements

The aim of this paper is to provide an alternative and more efficient screening test for allergy with respect to traditional UNICAP and prick skin tests, which are commercially available nowadays. The proposed test is aimed at monitoring the interaction of the IgE protein with the allergen in an allergic subject. The idea relies on the fact that the interaction between the IgE and the corresponding allergen happens through binding mechanisms between the IgE and the allergen structure, changing the ionic content of the serum. This latter can be monitored by impedance measurements and confirmed by reactive oxygen species (ROS) evaluation in leukocyte subpopulations. Therefore, in this paper we propose a novel system for the detection of allergies based on impedance measurements and confirmed by ROS measurements. This type of measurement approach has never been presented in the literature for this purpose. A measurement protocol has been developed to guarantee reproducibility and repeatability of the assay. This paper describes both the instrument—specifically designed for impedance evaluation—and the measurement protocols for both methodologies, defining univocally, in this way, the overall measurement technique.

New Insights into Cockroach Allergens.

This review addresses the most recent developments on cockroach allergen research in relation to allergic diseases, especially asthma. The number of allergens relevant to cockroach allergy has recently expanded considerably up to 12 groups. New X-ray crystal structures of allergens from groups 1, 2, and 5 revealed interesting features with implications for allergen standardization, sensitization, diagnosis, and therapy. Cockroach allergy is strongly associated with asthma particularly among children and young adults living in inner-city environments, posing challenges for disease control. Environmental interventions targeted at reducing cockroach allergen exposure have provided conflicting results. Immunotherapy may be a way to modify the natural history of cockroach allergy and decrease symptoms and asthma severity among sensitized and exposed individuals. The new information on cockroach allergens is important for the assessment of allergen markers of exposure and disease, and for the design of immunotherapy trials.

Immunotherapy for Food Allergy: Are We There Yet?

Current clinical research focuses on food allergen-specific immunotherapy through oral (OIT), sublingual (SLIT), or epicutaneous (EPIT) routes. Immunotherapy relies on the delivery of gradually increasing doses of specific allergens to induce desensitization (defined as temporary antigen hyporesponsiveness that depends on regular food ingestion) and, ultimately, tolerance (defined as the ability to ingest food without symptoms despite prolonged periods of avoidance or irregular intake). Although the majority of the patients treated with OIT achieve desensitization, only a minority achieves tolerance. OIT involves higher maintenance doses of food protein (300 mg-4g) compared with SLIT (2.5-7.5 mg) and EPIT (250-500 mcg). OIT efficacy is higher compared with SLIT, but OIT is associated with higher rate of systemic adverse events compared with SLIT and EPIT. OIT is also associated with a minor risk of eosinophilic esophagitis. Combined treatment of OIT and anti-IgE monoclonal antibody has improved safety but not efficacy compared with OIT alone. Early initiation of peanut OIT in peanut-allergic infants and young children may afford superior efficacy and safety. In this review, we discuss the allergen-specific strategies currently explored for the treatment of food allergy, including immunotherapy with native and heat-modified food proteins. Additional research employs strategies to improve the safety and efficacy of allergen immunotherapy through modifications of allergen structure and addition of immunomodulatory adjuvants.

Cross-React: a new structural bioinformatics method for predicting allergen cross-reactivity.

: Cross-React is available at: http://curie.utmb.edu/Cross-React.html. ssnegi@utmb.edu.

NMR resonance assignments of the major apple allergen Mal d 1.

The major apple allergen Mal d 1 is the predominant cause of apple (Malus domestica) allergies in large parts of Europe and Northern America. Allergic reactions against this 17.5 kDa protein are the consequence of initial sensitization to the structurally homologous major allergen from birch pollen, Bet v 1. Consumption of apples can subsequently provoke immunologic cross-reactivity of Bet v 1-specific antibodies with Mal d 1 and trigger severe oral allergic syndroms, affecting more than 70 % of all individuals that are sensitized to birch pollen. While the accumulated immunological data suggest that Mal d 1 has a three-dimensional fold that is similar to Bet v 1, experimental structural data for this protein are not available to date. In a first step towards structural characterization of Mal d 1, backbone and side chain 1H, 13C and 15N chemical shifts of the isoform Mal d 1.0101 were assigned. The NMR-chemical shift data show that this protein is composed of seven β-strands and three α-helices, which is in accordance with the reported secondary structure of the major birch pollen allergen, indicating that Mal d 1 and Bet v 1 indeed have similar three-dimensional folds. The next stage in the characterization of Mal d 1 will be to utilize these resonance assignments in solving the solution structure of this protein.

Development and Evaluation of Standards for Assessing the Biological Activity of Allergens

The guidelines for construction and approval of international standards and reference reagents of biological substances according to which allergens, like all immunobiological drugs, should be standardized were described. The units of biological (allergic) activity for the first batch of standard are determined in vivo (provocative tests, intradermal tests, skin-prick test, etc.) in sensitized persons. The activity of the first batch of the standard is also studied in parallel by immunoassay methods using inhibition of serum containing allergen-specific IgE. The standard should also be characterized with respect to content of main (major) allergenic proteins, protein profile, and protein structure. Modern requirements for allergen preparations were introduced for the first time in the XIIIth Edition of the State Pharmacopoeia of the Russian Federation.

Structural and Functional Characterization of the Hazelnut Allergen Cor a 8.

Nonspecific lipid transfer proteins (nsLTPs) are basic proteins, stabilized by four disulfide bonds, and are expressed throughout the plant kingdom. These proteins are also known as important allergens in fruits and tree nuts. In this study, the nsLTP from hazelnuts, Cor a 8, was purified and its crystal structure determined. The protein is stable at low pH and refolds after thermal denaturation. Molecular dynamics simulations were used to provide an insight into conformational changes of Cor a 8 upon ligand binding. When known epitope areas from Pru p 3 were compared to those of Cor a 8, differences were obvious, which may contribute to limited cross-reactivity between peach and hazelnut allergens. Differences in epitope regions may contribute to limited cross-reactivity between Cor a 8 and nsLTPs from other plant sources. The structure of Cor a 8 represents the first resolved structure of a hazelnut allergen.

Allergens in veterinary medicine.

Allergic diseases in animals are increasingly gaining importance in veterinary practice and as research models. For intradermal testing and allergen immunotherapy, a good knowledge of relevant allergens for the individual species is of great importance. Currently, the knowledge about relevant veterinary allergens is based on sensitization rates identified by intradermal testing or serum testing for allergen‐specific IgE; crude extracts are the basis for most evaluations. Only a few studies provide evidence about the molecular structure of (particularly) dust mite, insect and mould allergens in dogs and horses, respectively. In those species, some major allergens differ from those in humans. This position paper summarizes the current knowledge about relevant allergens in dogs, cats and horses.

К вопросу о разработке стандартных образцов биологической активности аллергенов и методах оценки их активности

Описаны принципы изготовления и утверждения международных стандартов и референс-реагентов биологических субстанций, согласно которым аллергены, как и все иммунобиологические лекарственные препараты, должны быть стандартизированы. Единицы биологической (аллергенной) активности для первой серии стандартного образца (СО) устанавливаются по in vivo методике (провокационные тесты, внутрикожные пробы, прик-тест и т.п.) на сенсибилизированных лицах. Активность первой серии стандарта также параллельно оценивается методами иммуноанализа с этапом ингибиции IgE-содержащих специфических сывороток к исследуемому аллергену. Помимо оценки биологической активности, СО должен быть охарактеризован по содержанию главных (мажорных) аллергенов: белку, белковому профилю и структуре белка. Современные требования, предъявляемые к препаратам аллергенов, впервые введены в XIII редакцию Государственной фармакопеи Российской Федерации.

The crystal structure of a common allergen in complex with its specific patient-derived antibody

The crystal structure of a common allergen in complex with its specific patient-derived antibody

Position paper of the EAACI: food allergy due to immunological cross-reactions with common inhalant allergens.

In older children, adolescents, and adults, a substantial part of all IgE-mediated food allergies is caused by cross-reacting allergenic structures shared by inhalants and foods. IgE stimulated by a cross-reactive inhalant allergen can result in diverse patterns of allergic reactions to various foods. Local, mild, or severe systemic reactions may occur already after the first consumption of a food containing a cross-reactive allergen. In clinical practice, clinically relevant sensitizations are elucidated by skin prick testing or by the determination of specific IgE invitro. Component-resolved diagnosis may help to reach a diagnosis and may predict the risk of a systemic reaction. Allergy needs to be confirmed in cases of unclear history by oral challenge tests. The therapeutic potential of allergen immunotherapy with inhalant allergens in pollen-related food allergy is not clear, and more placebo-controlled studies are needed. As we are facing an increasing incidence of pollen allergies, a shift in sensitization patterns and changes in nutritional habits, and the occurrence of new, so far unknown allergies due to cross-reactions are expected.

Analysis of glutathione S-transferase allergen cross-reactivity in a North American population: Relevance for molecular diagnosis

It is not clear whether cross-reactivity or cosensitization to glutathione S-transferases (GSTs) occurs in tropical and subtropical environments. In the United States, Bla g 5 is the most important GST allergen and lack of coexposure to GSTs from certain species allows a better assessment of cross-reactivity. To examine the molecular structure of GST allergens from cockroach (Bla g 5), dust mites (Der p 8 and Blo t 8), and helminth (Asc s 13) for potential cross-reactive sites, and to assess the IgE cross-reactivity of sensitized patients from a temperate climate for these allergens for molecular diagnostic purposes. Four crystal structures were determined. Sera from patients allergic to cockroach and mite were tested for IgE reactivity to these GSTs. A panel of 6 murine anti-Bla g 5 mAb was assessed for cross-reactivity with the other 3 GSTs using antibody binding assays. Comparisons of the allergen structures, formed by 2-domain monomers that dimerize, revealed few contiguous regions of similar exposed residues, rendering cross-reactivity unlikely. Accordingly, anti-Bla g 5 or anti-Der p 8 IgE from North American patients did not recognize Der p 8 or Bla g 5, respectively, and neither showed binding to Blo t 8 or Asc s 13. A weaker binding of anti-Bla g 5 IgE to Der p 8 versus Bla g 5 (∼ 100-fold) was observed by inhibition assays, similar to a weak recognition of Der p 8 by anti-Bla g 5 mAb. Patients from tropical Colombia had IgE to all 4 GSTs. The lack of significant IgE cross-reactivity among the 4 GSTs is in agreement with the low shared amino acid identity at the molecular surface. Each GST is needed for accurate molecular diagnosis in different geographic areas.

Interfaces between allergen structure and diagnosis: know your epitopes.

Allergy diagnosis is based on the patient's clinical history and can be strengthened by tests that confirm the origin of sensitization. In the past 25years, these tests have evolved from the exclusive in vivo or in vitro use of allergen extracts, to complementary molecular-based diagnostics that rely on in vitro measurements of IgE reactivity to individual allergens. For this to occur, an increase in our understanding of the molecular structure of allergens, largely due to the development of technologies such as molecular cloning and expression of recombinant allergens, X-ray crystallography, or nuclear magnetic resonance (NMR), has been essential. New in vitro microarray or multiplex systems are now available to measure IgE against a selected panel of purified natural or recombinant allergens. The determination of the three-dimensional structure of allergens has facilitated detailed molecular studies, including the analysis of antigenic determinants for diagnostic purposes.

Food Allergies: The Basics

IgE-associated food allergy affects approximately 3% of the population and has severe effects on the daily life of patients—manifestations occur not only in the gastrointestinal tract but also affect other organ systems. Birth cohort studies have shown that allergic sensitization to food allergens develops early in childhood. Mechanisms of pathogenesis include cross-linking of mast cell– and basophil-bound IgE and immediate release of inflammatory mediators, as well as late-phase and chronic allergic inflammation, resulting from T-cell, basophil, and eosinophil activation. Researchers have begun to characterize the molecular features of food allergens and have developed chip-based assays for multiple allergens. These have provided information about cross-reactivity among different sources of food allergens, identified disease-causing food allergens, and helped us to estimate the severity and types of allergic reactions in patients. Importantly, learning about the structure of disease-causing food allergens has allowed researchers to engineer synthetic and recombinant vaccines.

Known Allergen Structures Predict Schistosoma mansoni IgE-Binding Antigens in Human Infection.

The IgE response has been associated with both allergic reactions and immunity to metazoan parasites. Recently, we hypothesized that all environmental allergens bear structural homology to IgE-binding antigens from metazoan parasites and that this homology defines the relatively small number of protein families containing allergenic targets. In this study, known allergen structures (Pfam domains) from major environmental allergen families were used to predict allergen-like (SmProfilin, SmVAL-6, SmLipocalin, SmHSP20, Sm triosephosphate isomerase, SmThioredoxin, Sm superoxide dismutase, SmCyclophilin, and Sm phosphoglycerate kinase) and non-allergen-like [Sm dynein light chain (SmDLC), SmAldolase SmAK, SmUbiquitin, and Sm14-3-3] proteins in Schistosoma mansoni. Recombinant antigens were produced in Escherichia coli and IgG1, IgG4, and IgE responses against them measured in a cohort of people (n = 222) infected with S. mansoni. All allergen-like antigens were targeted by IgE responses in infected subjects, whilst IgE responses to the non-allergen-like antigens, SmAK, SmUbiquitin, and Sm14-3-3 were essentially absent being of both low prevalence and magnitude. Two new IgE-binding Pfam domain families, not previously described in allergen family databases, were also found, with prevalent IgE responses against SmDLC (PF01221) and SmAldolase (PF00274). Finally, it was demonstrated that immunoregulatory serological processes typically associated with allergens also occurred in responses to allergen-like proteins in S. mansoni infections, including the production of IgG4 in people responding with IgE and the down-regulation of IgE in response to increased antigen exposure from S. mansoni eggs. This study establishes that structures of known allergens can be used to predict IgE responses against homologous parasite allergen-like molecules (parallergens) and that serological responses with IgE/IgG4 to parallergens mirror those seen against allergens, supporting our hypothesis that allergenicity is rooted in expression of certain protein domain families in metazoan parasites.